This application will characterize how age decreases the ability of the normal tissue microenvironment to regulate cellular differentiation/function, suppressing expression of the tumorigenic phenotype. This project will utilize a unique experimental system in which rat liver epithelial tumor cells that carry a transfected bacterial beta- galactosidase reporter gene are transplanted into the liver parenchymal or subcutaneous tissue compartments of syngeneic rats. The ability of the normal hepatic parenchyma to suppress the tumorigenic potential of transplanted, neoplastically transformed rat liver epithelial cells can be evaluated directly using beta-galactosidase histochemistry. We have previously characterized the transplantation site-specific tumorigenic potential of two rat liver epithelial tumor cell lines. In the young adult rat (3-months old), these cell lines form undifferentiated spindle-cell carcinomas with short latency at subcutaneous transplantation sites, but in the liver form slow-growing hepatocellular carcinomas or no tumors at all. Suppressed tumor cells incorporate into hepatic plates and morphologically differentiate into hepatocyte-like cells. The tumor cell line that is suppressed in the livers of young adult males forms undifferentiated carcinomas in the livers of 100% of 24-month old rats. These results demonstrate that age of host and transplantation site are strong determinants in the tumorigenic potential of transplanted tumor cells. We hypothesize that the liver microenvironment exerts a strong regulatory (suppressive) capacity over transplanted cells and that this capacity-is lost With age. The transplantation site-specificity of tumor suppression was evidenced by tumor formation at extrahepatic sites of animals that formed no hepatic tumors following transplantation of cells directly into the hepatic parenchyma. The first goal of this project is to extend our observations on the loss of liver-specific tumor suppression in older male rats to a full range of ages of recipient rats, and to evaluate the hormonal component of the male predominance of liver cancer using female and castrated male rats of various ages. The second goal is to investigate the role of Kupffer and pit cells on hepatic tumor formation by examining their numbers/distribution in vivo. We will also evaluate the effects of growth modulating factors produced by these cells on tumor cell proliferation/differentiation in vitro. The third goal is to develop an in vitro system for evaluating the interaction of tumor cells with specific extracellular matrix and cell membrane components, and .humoral factors, isolated from rats of different ages and gender. The final goal is to characterize, using the technique of differential display, changes in gene expression patterns that accompany the transplantation of tumor cells into different tissue microenvironments.